Integrins regulate cell growth, differentiation, and behavior in many systems. Integrin β_1C (β_1S) is an alternatively spliced variant of integrin β₁ with a specific cytoplasmic domain and is expressed in several human tissues. Human β_1C transiently expressed in mouse 10T1/2 fibroblasts showed a diffuse pattern of cell surface staining, whereas β₁ localized to focal adhesions. Moderate concentrations of β_1Chad no effect on actin stress fibers or focal adhesions, but markedly inhibited DNA synthesis. Inhibition by β_1C mapped to the late G₁ phase of the cell cycle, near the G₁-S boundary. Thus, alternative splicing of β₁ results in transmission of distinct signals that may regulate growth in vivo.